Infusion Of CAR Research Articles

Resolution of Anterior Uveal Infiltration of Acute Lymphoblastic Leukemia After Chimeric Antigen Receptor T-Cell Therapy

Purpose: To present a case of acute B-cell lymphoblastic leukemia (B-ALL) ocular relapse treated with chimeric antigen receptor T-cell (CAR T) therapy. Methods: A retrospective case review was evaluated. Results: A 69-year-old woman with a history of B-ALL presented with bilateral hypopyons and 3+ anterior chamber cells that responded to topical prednisolone. Six months later, she developed recurrent hypopyons, iris neovascularization, and iris nodularity in both eyes. A fine-needle aspiration biopsy confirmed ocular infiltration of B-ALL. A single infusion of CAR T therapy resulted in remission of the systemic and ocular findings. The clinical course was complicated by cytokine release syndrome and neurotoxicity, which improved with medical management. The patient remained in remission for 7 months after a single CAR T infusion. Conclusions: CAR T therapy may be effective in treating systemic leukemia and uveal infiltration, with an ocular side-effect profile and visual outcomes. The systemic side effects of CAR T therapy may be managed medically.

Evaluation of all-cause mortality and cardiovascular safety in patients receiving chimeric antigen receptor T cell therapy: a prospective cohort study

Assessment of cardiovascular risk is critical for patients with cancer. Previous retrospective studies suggest potential cardiotoxicity of CAR T cell therapies. We aimed to prospectively assess cardiotoxicity and the predictive value of cardiac biomarkers and classical risk factors (age, cardiac function, diabetes, arterial hypertension, smoking) for cardiac events and all-cause mortality (ACM). In this prospective cohort study, all patients treated with CAR T cell constructs (axi-cel, tisa-cel, brexu-cel, ide-cel, or the 3rd generation CAR HD-CAR-1) from Oct 1, 2018, to Sept 30, 2022at the University Hospital Heidelberg were included. Surveillance included cardiac assessment with biomarkers (high-sensitive Troponin T (hs-cTnT), N-terminal brain natriuretic peptide (NT-proBNP)), 12-lead-ECG, and 2D echocardiography. ACM was defined as the primary study endpoint, while cardiotoxicity, defined by clinical syndromes of heart failure or decline in ejection fraction, served as a secondary endpoint. Overall, 137 patients (median age 60, range 20-83, IQR 16), were included in the study. 46 patients died during the follow up period (median 0.75 years, range 0.02-4.33, IQR 0.89) 57 month, with a median survival of 0.57 years (range 0.03-2.38 years, IQR 0.79). A septal wall thickness above 11mm (HR 2.48, 95%-CI=1.10-5.67, p=0.029) was associated with an increased risk of ACM, with a trend seen for reduced left ventricular ejection fraction prior to therapy (LVEF <40%; HR 9.17, 95%-CI=1.30-183.11, p=0.051). Secondary endpoint was reached by 93 patients while no baseline parameter was able to predict an elevated risk. However, hs-cTnT change from baseline of 50% or more during the first 14 days after CAR infusion predicted ACM (HR 3.81, 95%-CI=1.58-9.45; p=0.003). None of the baseline characteristics was able to predict the incidence of cardiac events. Reduced pre-lymphodepletion ejection fraction and early post-infusion biomarker kinetics may be associated with increased ACM and cardiotoxicity events. These findings may help to identify patients who could benefit from intensified cardio-oncological surveillance. The German Center for Cardiovascular Research, German Research Foundation, and the Federal Ministry of Education and Research.

A Phase II Trial of Prophylactic Anakinra to Prevent Neurotoxicity in Patients Receiving Anti-CD19 CAR T-Cell Therapy for Relapsed or Refractory Lymphoma: Final Results from Cohort 2

Background: Whilst tocilizumab is effective in the management of chimeric antigen receptor T-cell (CAR-T) associated cytokine release syndrome (CRS), there are no targeted therapies for immune effector cell-associated neurotoxicity syndrome (ICANS). Rates of severe (≥ grade 3) ICANS remain high with axicabtagene (axi-cel) and brexucabtagene (brexu-cel) at &amp;gt; 30%. Preclinical models have shown that anakinra, an IL-1 receptor antagonist, reduced CAR-mediated immune toxicities without affecting efficacy. Based on these data, we conducted a phase 2 trial of prophylactic anakinra in patients receiving commercial anti-CD19 CAR-T for r/r B-cell lymphoma (NCT04148430). The study had sequential enrollment in 2 independent cohorts. We recently reported results from cohort 1 in which patients received subcutaneous (SC) anakinra from day 2 until at least day 10 post-CAR infusion and showed that prophylactic anakinra reduced the rate of severe ICANS to 9.7% (Park, Nat Med, 2023). Herein, we report results from cohort 2 of the study where we utilized anakinra as a complete prophylactic strategy. Methods: Adult patients with a diagnosis of r/r large B-cell lymphoma (LBCL), high-grade B-cell lymphoma (HGBCL), follicular lymphoma (FL) or mantle cell lymphoma (MCL) receiving commercial anti-CD19 CAR-T were enrolled. Patients received SC anakinra 100 mg daily from day 0 for at least 7 d. The frequency of anakinra could be increased up to 100 mg every 6 h, and the duration beyond 7 d for persistent or progressive CRS and ICANS. Tocilizumab and steroid use were allowed for the management of persistent or progressive CRS and ICANS. A baseline and day 5 lumbar puncture were performed for CSF analysis. The primary endpoint was the rate of severe ICANS, graded per the ASTCT criteria. Secondary endpoints included rates of severe CRS, all grade CRS and ICANS, overall response rate (ORR), survival outcomes, and serum and CSF biomarker levels. Results: 37 patients were screened, 31 enrolled, and 30 treated (LBCL=13, HGBCL=6, FL=6, MCL=5). 1 enrolled patient did not proceed with treatment due to COVID-19 infection. The median age of the cohort was 65yrs (range: 24 - 81yrs) and patients received axi-cel (19 pts; 63%), brexu-cel (5 pts; 17%), lisocabtagene (4 pts; 13%) and tisagenlecleucel (2 pts; 7%). Patients received a median 3 prior lines of therapy (range: 1 - 7); 22 (73%) were refractory to their immediate prior therapy; 17 (57%) received bridging therapy and 3 (10%) had active CNS disease at CAR infusion. Pretherapy tumor burden as measured by the median sum of the products of diameters was 1638mm 2 (range: 0 - 21,570mm 2). All patients started anakinra on day 0. Anakinra administration was increased in 24 patients (80%) during CRS; it was increased to a maximum of 100mg q12h in 1 patient and 100mg q6h in 23 patients. The median duration of anakinra administration was 9.5 d (IQR 8 - 13). No patient stopped anakinra due to an adverse event. A total of 3 patients (10%) experienced severe ICANS, all of which were grade 3 only. 9 patients (30%) experienced any grade ICANS (Fig 1A). The median time to ICANS onset was 6.5 d (range: 4 - 10 d). All-grade CRS occurred in 24 patients (80%). Most CRS events were grade 1 or 2 except for 3 grade 3 events (10%) (Fig 1A). Of the 24 patients who received CD28-containing CARs (axi-cel and brexu-cel), all-grade and severe ICANS rate was 29% and 12.5%, respectively (Fig 1B). In the 3 patients with active CNS disease, there was 1 grade 2, and 1 grade 3 ICANS event. 23 patients (77%) received tocilizumab and 18 (60%) received steroids for CRS and/or ICANS. The median follow-up was 5.8mo. The best ORR by day 100 was 83% (95% CI 65-94%) with a CR rate of 73% (95% CI 54-89%). The estimated progression-free and overall survival at 6mo was 72% (95% CI 50-93%) and 81% (95% CI 66-86%), respectively. Conclusion: Early use of prophylactic anakinra was safe, feasible and reduced the rate of severe ICANS without affecting efficacy in adult patients receiving anti-CD19 CAR-T for r/r B-cell lymphoma. Rates of severe ICANS were very similar between cohorts 1 and 2; at 9.7% and 10% in the entire cohorts, respectively, and 11% and 12.5% in patients treated with CD28-containing CARs, respectively. The risk-adapted dosing of anakinra likely contributed to the low rates of severe ICANS in this study. Detailed comparison of results from cohort 1, analysis of the anakinra dose-relationship on efficacy, and serum and CSF biomarkers will be presented at the meeting.

Real-Word Experience of CAR T-Cells in Patients with Relapsed/Refractory Follicular Lymphoma : A Descart Registry Analysis from the Lysa

Background: Anti-CD19 CAR T-cells have revolutionized the treatment of aggressive B-cell non-Hodgkin lymphomas (NHL) by demonstrating durable responses. Although follow-up remain short, axicabtagene ciloleucel (axi-cel, ZUMA-5) and tisagenlecleucel (tisa-cel, ELARA) demonstrated promising complete response rates (CRR) of 86% and 69%, respectively, in phase II trials including relapsed or refractory (R/R) follicular lymphoma (FL).. Tisa-cel and axi-cel are now approved after at least 2 lines of systemic therapy by the FDA. Few real-word evidence (RWE) data have been reported so far. Methods: Patients were eligible if aged &amp;gt;18 years, had FL histology (grade 1-3A) and had received at least 2 previous treatment lines for tisa-cel and 3 for axi-cel according to the French early access program label. From Dec 2021 to Jan 2023, a total of 112 patients were included in early access program by 21 French centers and in DESCAR-T registry (NCT04328298), among them 87 were infused with a CAR product. At the data cut-off on March 2023, 70 had at least 1 month of follow-up (FU) with PET-CT evaluation, and thus were considered for the safety and efficacy sets (62 tisa-cel and 8 axi-cel,). Results: Patient and disease characteristics were as follows: 46 males (65.7%), median age 62 years (range: 34-79), 3 median prior lines of therapy (range 2-9, including bi-specific antibody in 12.9% and autoSCT in 44.3%), FLIPI 0-2 in 40.5% and 3-5 in 49.5%, bulky disease (&amp;gt;5cm) in 22%, and LDH &amp;gt; N 52.2%. POD24 after 1 st systemic immunochemotherapy (IC) was reported in 44 patients (62.8%). Before CAR T-cell infusion, 58.6% of patients received a bridging therapy (20 (48.8%) chemotherapy, 25 (61%) monoclonal antibodies, 4 (9.8%) kinase inhibitors, 15 (36.6%) lenalidomide, and 3 (7.3%) radiotherapy). All patients but one received a fludarabine and cyclophosphamide-based lymphodepletion. Median time from order to infusion was 48 days (range 34-204), and 41 days (30-328) from leukapheresis to infusion. Median FU was 7.3 months [6.4; 8.2] from product order, and 5.4 months [3.4; 6] from CAR infusion. Best ORR and CRR were 97.5 and 87.5%, respectively with 72.5% of patients in CR at 1 month with projected DOR and DOCR of 72.4% [56.8; 83.1%], and 79.7% [62.7%; 89.6%], respectively. Projected 6-months PFS, OS, were 71.8% [95% CI, 56.6%; 82.4%], 97.4% [83.2%; 99.6%], respectively. Fifteen (21.4%) patients progressed (median time from infusion to first relapse: 3 months (range 1-10 mo). Only one patient died from lymphoma progression. Any-grade CRS and ICANS were reported for 74.3% and 27.1% of patients, but grade 3-4 CRS/ICANS were very unfrequent,1.4% and 4.3%, respectively. No grade 5 was reported. Persisting grade 3-4 hematologic toxicities at 1 and 3 months were: neutropenia (50% and 12.3%), thrombocytopenia (18.6% and 0%), and anemia (8.6% and 0%). Medically relevant bacterial and viral infections were reported in 20% and 14.3% of patients, respectively. Conclusions: Although longer FU is needed to assess disease control, RWE data from the DESCAR-T registry confirm the excellent response rates and safety profile of CAR T-cells in R/R FL after at least 2 lines of previous therapy.

Trial-in-Progress: A Phase 1/2 Multi-Center Study of Onct-808, a ROR1-Specific CAR T, in Adult Patients with Relapsed/Refractory Aggressive B Cell Lymphoma

BACKGROUND: The therapeutic landscape for B-cell lymphomas (BCL) leverages several modalities, including chemotherapy, targeted agents, adoptive cell therapy, and immunotherapy. While these approaches have shown clinical benefit, challenges remain, such as variable response rates, treatment resistance, and toxicities. ROR1-specific CAR T-cell therapy represents a promising potential advancement in the treatment of aggressive BCL in patients (pts) who have failed previous CD19 CAR T therapy. ROR1-targeted CAR T therapy offers several advantages that could address these challenges. By directing CAR T cells to specifically recognize and eliminate ROR1-expressing tumor cells, it offers a highly targeted approach with the potential for enhanced efficacy. Moreover, ROR1 is often highly expressed on aggressive BCL, making it a suitable therapeutic target. This focused approach may help overcome the limitations associated with current treatments and improve response rates. After birth ROR1 is predominantly expressed on malignant cells; therefore, this selective expression pattern may reduce the risk of toxicities, including off-tumor target elimination and non-specific activation of immune cells. The development of ROR1-specific CAR T holds promise in enhancing the treatment outcomes for patients with aggressive BCL. With targeted action and potential to reduce toxicity, ROR1-specific CAR T cell therapy offers an innovative and novel strategy for improving patient outcomes in BCL. The ROR1 binding moiety for ONCT-808 is derived from zilovertamab. Zilovertamab vedotin showed preliminary evidence of efficacy and no evidence of on-target off-tumor toxicity in patients with advanced B cell malignancies (Wang 2022). METHODS: This is a Phase 1/2, single-arm, open-label, multi-center trial to evaluate the safety and efficacy of ONCT-808 in pts with aggressive BCL, including large B-cell lymphoma (LBCL) and mantle cell lymphoma (MCL) with no approved therapy available. The study comprises two distinct phases: Phase 1 for dose escalation followed by Phase 2 for dose expansion. In Phase 1, a standard 3+3 dose escalation design will be used with 3 provisional dose levels and expected sample size of up to 18 patients. Subjects will receive a conditioning regimen of intravenous (IV) cyclophosphamide and fludarabine, followed by ONCT-808 IV infusion. The dose of ONCT-808 will be sequentially escalated to determine the recommended Phase 2 dose (RP2D) for LBCL and MCL cohorts. Bridging therapy is permitted if clinically indicated. Phase 2 will enroll LBCL and MCL patients into separate dose expansion cohorts at the RP2D determined in Phase 1. ONCT-808 will be administered as a single IV infusion of autologous CAR T cells. The primary outcomes include assessing the incidence, severity, and relationship of Dose Limiting Toxicities (DLT), Treatment Emergent Adverse Events (TEAE), and RP2D selection of ONCT-808. Secondary outcomes include evaluating the Overall Response Rate (ORR), Complete Response (CR) rate, Duration of Response (DOR), pharmacokinetics of ONCT-808, expansion, persistence of ROR1 CAR-positive T cells, overall safety and tolerability. Key inclusion criteria include histologically confirmed aggressive B-cell NHL, measurable lesions per Lugano criteria (Cheson, 2014), and relapsed/refractory (R/R) disease without available therapy (must be previously treated with systemic therapy, progressed after or ineligible for autologous HSCT, and received prior approved anti-CD19 CAR-T at least 6 months prior to enrollment unless refused/ineligible). Key exclusion criteria include previous ROR1-targeted therapy, ongoing immunosuppression, and CNS involvement. The trial is currently active and patients have been treated in the phase 1 dose escalation. Clinical Trial Identifier: NCT05588440

A Phase I Study of CD19-Targeted 19(T2)28z1xx CAR T Cells in Adult Patients with Relapsed or Refractory Diffuse Large B-Cell Lymphoma

Background: Autologous CD19 CAR T cell therapies have demonstrated favorable clinical responses in relapsed or refractory (R/R) diffuse large B-cell lymphomas (DLBCL), but only a subset of patients (pts) experience durable remissions. We hypothesized that the redundancy of CD28 and CD3ζ signaling in a CAR design incorporating all 3 CD3ζ immunoreceptor tyrosine-based activation motifs (ITAMs) might negatively affect T cell differentiation and promote exhaustion. Therefore, we created a new CD19 CAR construct with calibrated CAR activation potential by mutating 2 of the 3 ITAMs, termed 1XX, and hypothesized that a 1XX CAR would afford durable responses at lower dose and fewer toxicities compared to CARs comprising 3 ITAMs. We previously presented the data from the dose-escalation portion of the phase I clinical trial of 19(T2)28z-1XX CAR T cells in DLBCL (Park J et al. ASH 2022). We have now completed accrual and report the updated results from both dose escalation and expansion portion of the study with a longer follow-up (NCT04464200). Methods: This is a single-center, phase I, dose-escalation and expansion trial of 19(T2)28z1XX CAR T cells in adult pts with R/R DLBCL conducted at Memorial Sloan Kettering Cancer Center. Pts received fludarabine and cyclophosphamide lymphodepleting chemotherapy followed by a single infusion of CAR T cells. There were 4 dose levels in the dose escalation cohort: dose level (DL) 1 (25x10 6), DL2 (50x10 6), DL3 (100x10 6) and DL4 (200x10 6 CAR T cells). The primary objective of the trial was to evaluate safety and tolerability and determine the recommended phase 2 dose. Key secondary objectives include assessment of overall response and complete response rates. Results: 28 pts have been enrolled and treated; 16 pts on the dose escalation and 12 pts on the dose expansion phase of the study. Median age was 62 (range, 48-86). 20 pts (71%) had primary refractory disease, 3 pts (11%) had prior autologous HSCT, and 2 pts (7%) had prior autologous CD19 CAR T therapy. In the dose-escalation phase (n=16), 15 pts experienced grade 1-2 CRS (94%) and no pt experienced ≥grade 3 CRS. Two pts (13%) experienced ICANS: 1 pt with grade 1 and 1 pt with grade 3 (Table 1). Eight pts received tocilizumab and 5 pts received corticosteroid. Complete response (CR) was observed in 11 of 16 pts (69%) and partial remission (PR) in 1 pt (6%) with overall response rate (ORR) of 75%. Responses were seen across all dose levels (Table 1). The lowest dose (DL1: 25x10 6) was selected for dose-expansion based on exposure-response analysis and 12 additional pts were treated at that dose level. In a pooled analysis of 16 pts who received DL1 dose (4 in dose escalation and 12 in dose expansion), 12 pts and 1 pt experienced grade 1-2 (75%) and grade 3 CRS (6%), respectively. Only 1 pt experienced ICANS (6%) at grade 3 (same pt who had grade 3 CRS), which was reversible and transient (Table 1). 11 pts received tocilizumab and 9 pts received corticosteroids. Twelve pts achieved CR (75%) and 2 pts (12%) PR with the best ORR of 87% (Table 1). A lower metabolic tumor volume (MTV) at screening but not at post-bridging was associated with a higher likelihood of achieving CR (p=0.039). With a median follow-up of 345 days (range, 33-1024), 4 pts experienced relapse (1 CR and 3 PR pts) and 16 pts have ongoing CR &amp;gt; 6 months (Figure 1). While CAR T cell persistence analysis is ongoing, 5 pts have CAR T cells detected beyond one year in ongoing CR including a patient treated at DL1 with detectable CAR T cells at 2 years. Conclusions: Treatment with 19(T2)28z1XX CAR T cells was shown to be safe with durable remissions with low rates of severe CRS (4%) and ICANS (7%). The overall CR rate of 71% is encouraging and appears durable with the majority of CR patients in ongoing remission &amp;gt; 6 months at a median follow-up of 345 days and with persisting CAR T cells. With more patients treated at DL1 with a longer follow-up, the study demonstrates a high efficacy of 19(T2)28z1XX CAR T cells at a dose significantly lower (25x10 6) than those of approved CD19 CAR T cells (100-300x10 6) and support that the 1XX single ITAM signaling domain may lead to enhanced efficacy of CD19 CARs and allow infusion of a lower T cell dose with favorable toxicity profiles.

A Comparative Analysis of Infectious Complications in Patients with Multiple Myeloma Treated with BCMA-Targeted Bispecific Antibodies and CAR T-Cell Therapy

Introduction: There is limited data comparing infectious toxicities in patients with relapsed/refractory multiple myeloma (RRMM) treated with T-cell engaging therapies, which include chimeric antigen receptor T-cell therapy (CAR-T) and bispecific antibodies (BsAb). There are now three US FDA approved B-cell maturation antigen (BCMA)-targeting T-cell engagers, and several more in ongoing clinical development. Awareness regarding their infectious toxicities may inform treatment selection and mitigation strategies. Methods: We conducted a single-center, observational study in patients with RRMM comparing infectious complications in patients treated with a commercial or investigational autologous BCMA-targeting CAR-T versus patients treated with a commercial or investigational BCMA-targeting BsAb. The date of CAR infusion (day 0) was between 03/22/2017 - 02/27/2023 for the CAR-T patients. For the BsAb cohort, the date of treatment initiation (day 0) was between 01/21/2020 - 02/07/2023. All infection-specific variables were collected from day 0 until the date of next line of therapy or last follow-up, with a data cut-off of 06/01/23. Infectious events were graded according to CTCAE version 5.0. Prophylactic antimicrobials were administered according to institutional and protocol guidelines. The primary endpoint was the incidence of severe (grade ≥3) infections. Secondary objectives included the time to 1 st infection, infection rate over time, infectious organisms, the impact of prolonged cytopenias, and the utility of intravenous immunoglobulin (IVIg) administration in preventing infections. Results: Of the total 147 patients, there were 92 CAR-T and 55 BsAb treated patients. The median age of the CAR-T and BsAb cohort was 62yrs vs 65yrs, respectively (P=0.043). CAR-T patients had a median of 6.5 prior lines of therapy (IQR 5-8) versus 6.0 (IQR 4-9) in the BsAb cohort (P = 0.7); 97% of CAR-T patients had a prior autologous transplant compared to 75% in the BsAb cohort (P &amp;lt; 0.001). In keeping with current clinical practice, 18 patients (33%) in the BsAb cohort had prior exposure to CAR-T whilst no patients in the CAR-T cohort had prior BsAb exposure. The median follow-up duration for infectious events was similar in both groups at 5.8 months (IQR 3.8-9.2) for CAR-T and 4.3 months (IQR 3.2-9.8) for the BsAb cohort. A total of 209 infections were reported: 115 with CAR-T and 94 with BsAb. In the CAR-T cohort 24/92 patients (26%) experienced ≥1 severe infection, all of which were grade 3 and there were no grade 4 or 5 events. A numerically higher incidence was seen in the BsAb cohort with 21/55 BsAb patients (38%) experiencing ≥1 severe infection (P = 0.14). Nineteen BsAb patients (35%) experienced grade 3 infections, 2 (3.6%) had grade 4 infections and 4 (7.3%) had grade 5 infections. Six CAR-T (6.5%) and 10 BsAb patients (18.2%) had &amp;gt;1 grade 3 infection. The incidence of ≥1 severe infection remained high in the BsAb cohort at 16/37 (43%) even after excluding the 18 BsAb patients with prior CAR-T exposure. The median time from treatment initiation to the first infection of any-grade was 2.5 months (95% CI 1.2 - NR) with CAR-T compared to 3.4 months (95% CI 1.6 - 6.8) with BsAb (P = 0.6). The rate of any-grade infections was similar between the two groups early post-therapy (before day 100). However, there was a significantly higher infection rate of any-grade after day 100 with BsAb compared to CAR-T with a median of 0.24 (IQR 0.18 -0 .34) vs 0.13 (IQR 0.08 - 0.17) infections per 30 days, respectively (P &amp;lt; 0.001) (Figure 1A). Regarding the time to the first grade ≥3 infection, 79% occurred within day 100 for CAR-T patients compared to only 48% in the BsAb cohort (Figure 1B). The proportion of bacterial, viral, fungal, and parasitic infections in the CAR-T group was 49% (n = 56), 48% (n = 55), 4.3% (n = 5) and 0.9% (n = 1) respectively, and in the BsAb group was 51% (n = 48), 44% (n = 41), 3.2% (n = 3), and 0 respectively. Conclusion: Infectious complications were common early after BCMA-targeting BsAb and BCMA CAR-T and declined over time. In this real-world comparison, distinct from their CAR-T counterparts, BsAb recipients appeared to have a more persistent infection risk and higher incidence of severe infections, which included four patients having a grade 5 infection. Further analysis, including the impact of prolonged cytopenias and the utility of IVIg administration will be presented at the meeting.

Effect of bispecific B7H3 x CD19 CAR T cells on host CD19 expression and CAR T cell engraftment.

10043 Background: Our experience of B7H3 chimeric antigen receptor T cells (CAR-T) revealed modest engraftment (max 4.9 cells/uL) and no objective responses on first infusion. Previous CD19 CAR T cell trials have revealed that host B Cell CD19 can drive robust engraftment and persistence even in the absence of malignant CD19 expression. We therefore developed a bispecific B7H3xCD19 CAR T cell with the intent of using host CD19 expression to drive better engraftment of the B7H3-targeting CAR. Methods: Young patients with relapsed/refractory solid tumors (R/RST) were enrolled onto a Phase 1 trial (NCT04483778) to examine the safety of autologous T cells lentivirally transduced to express either or both scFV-IgG4hinge-CD28tm-4-1BB-zeta B7H3-specific and CD19-specific CARs. The B7H3 CAR construct contained the methotrexate resistance/selection cassette DHFRdm and the tracking/suicide construct EGFRt and the CD19 CAR construct containing the tracking/suicide construct HER2tg, allowing for differential CAR tracking. CAR T cells were cultured in low-dose methotrexate to select for B7H3 containing CAR T cells and limit exposure to monospecific CD19 CAR T cells. All patients received lymphodepleting fludarabine and cyclophosphamide prior to infusion of cryopreserved CAR-T at the prescribed dose level. The maximal tolerated dose or biologically effective dose (BED) was determined based upon observed toxicity through day 28 from initial CAR-T infusion and using a 3+3 statistical design. Results: 11 subjects (age 5-23, median 18 years) enrolled and received dose level (DL) 0B (0.5 x 106 TOTAL CAR-T/kg, n = 7) or DL1B (1 x 106 TOTAL CAR-T cells/kg, n = 4). One subject experienced dose limiting toxicities at DL1 of transaminitis, hyponatremia, worsening of a pericardial effusion and hypoxia. Most common toxicities were cytokine release syndrome (CRS) (n = 4, maximum CTCAE grade 2).No occurrence of ICANS was noted. Maximum total circulating CAR-T expansion on first infusion was 796.3 cells/uL and max B7H3 expansion was 374.2 cells/uL with 7/11 continuing with ongoing detection of CAR T cells in the peripheral blood at time of last follow up (21 days to 8 months). CAR engraftment was predominated by the B7H3 CAR single expressing CAR T cells in all subjects. All subjects developed B cell aplasia which is ongoing in 10/11 subjects at time of last check (range of 21 days to 8 months). Best overall response of Stable Disease was observed in 2 of the 11 subjects infused. Conclusions: B7H3xCD19 CAR T cells robustly engraft and expand in patients with R/RST and are safe, but despite ongoing detection of the CAR T cells, no objective responses were observed. It is likely that additional measures will be required to enhance the activity of CAR-T cells to achieve objective responses. STRIvE-02 Arm C will explore the addition of pembrolizumab post bispecific CAR infusion to evaluate the effect of PD1 blockade on anti-tumor activity. Clinical trial information: NCT04483778 .

Allogeneic CAR T Cells Targeting DLL3 Are Efficacious and Safe in Preclinical Models of Small Cell Lung Cancer.

Small cell lung cancer (SCLC) is an aggressive disease with limited treatment options. Delta-like ligand 3 (DLL3) is highly expressed on SCLC and several other types of neuroendocrine cancers, with limited normal tissue RNA expression in brain, pituitary, and testis, making it a promising CAR T-cell target for SCLC and other solid tumor indications. A large panel of anti-DLL3 scFv-based CARs were characterized for both in vitro and in vivo activity. To understand the potential for pituitary and brain toxicity, subcutaneous or intracranial tumors expressing DLL3 were implanted in mice and treated with mouse cross-reactive DLL3 CAR T cells. A subset of CARs demonstrated high sensitivity for targets with low DLL3 density and long-term killing potential in vitro. Infusion of DLL3 CAR T cells led to robust antitumor efficacy, including complete responses, in subcutaneous and systemic SCLC in vivo models. CAR T-cell infiltration into intermediate and posterior pituitary was detected, but no tissue damage in brain or pituitary was observed, and the hormone-secretion function of the pituitary was not ablated. In summary, the preclinical efficacy and safety data presented here support further evaluation of DLL3 CAR T cells as potential clinical candidates for the treatment of SCLC.

Distinguishing Early Infections from CRS with Routine and Exploratory Serum Proteomics and the HT10 Score Following CD19 CAR-T for Relapsed/Refractory B-NHL

Background: Early fever after CAR-T therapy can reflect both an infection or CRS. Distinguishing both etiologies in these commonly neutropenic patients represents an unresolved clinical challenge. Accordingly, the vast majority of patients receive empiric broad-spectrum antibiotics, which can promote antibiotic resistance and negatively influence the gut microbiome, which has recently been linked to inferior CAR-T outcomes (Smith et al, Nature Medicine 2022). Here, we aimed to characterize infection- vs. CRS-specific differences in routine and exploratory serum proteomics, and to develop a clinical tool that reliably discriminates between both. Methods: Early fever events (day 0-30) were retrospectively characterized as CRS vs. infection in 62 patients treated with standard-of-care CD19 CAR-T for r/r B-NHL (23 Axi-cel, 30 Tisa-cel, 9 Brexu-cel). Severe infections were defined as requiring IV anti-infectives and/or hospitalization in the presence of microbiologic evidence or distinct clinical signs of infection; CRS was graded according to ASTCT criteria. Cumulative incidence curves were calculated as time-to-event-analysis for CRS 1°, ≥2° and severe infections (day of event = index event). Serum inflammatory markers (CRP, interleukin-6 [IL-6], procalcitonin [PCT]) were measured daily from CAR infusion until discharge and on subsequent outpatient visits. Inflammatory markers were tested using binary logistic regression for severe infection vs. CRSonlyindex events. Discriminatory thresholds were determined from receiver operating characteristic (ROC) curves by optimizing the Youden J statistic. Longitudinal plasma proteomics was performed in 52 patients across four time points (day 0, 4, 14, 28) using a 92-protein multiplex proximity extension assay (Olink Proteomics). We applied a linear mixed model (LMM) accounting for both infection vs. CRS status and time effects. Results: The cumulative 30-day incidence of CRS 1°, CRS ≥2° and severe infections was 47%, 43%, and 25%, respectively. Five patients had neither CRS nor an infection, and were subsequently excluded from further analysis. Tocilizumab was applied in the majority of CRS patients (1°: 90%, ≥2°: 100%). We did not find a significant correlation between CRS grade and severe infections (G2 = 0.13, p = n.s.). Median onset of CRS and infection was 3 and 8 days, respectively. Most events (54%) occurred in the setting of coincident severe neutropenia (ANC <500/µl). While event-day CRP was comparable between infection and CRS (median 1.8 vs. 3.4 mg/dL, n.s.), we noted increased serum IL-6 (median 2243 vs. 64 pg/mL, p=0.03) and particularly high PCT levels (median 1.6 vs. 0.3 µg/L, p<0.0001) in the infection cohort. For PCT, an optimal discriminatory threshold of 1.5 µg/L was established (AUCROC 0.78). In previous work, we demonstrated that a CAR-HEMATOTOX (HT) score ≥2 identifies patients with a high infection risk (Rejeski et al, JITC 2022). Indeed, the infection cohort displayed higher baseline HT scores (median 5 vs. 2, p=0.002). By incorporating day-of-fever PCT levels with the HT score, the discriminatory capacity for severe infection events was improved. On ROC analysis, the 'HT10 score' (addition of 10 points to the patient-specific HT score in case event-day PCT ≥1.5 µg/L) exhibited superior discrimination for severe infections (AUC 0.92, p<0.0001, sens. 80%, spec. 91%, Fig. 1A). Additionally, we identified protein-level differences between the infection (n=13) vs. CRSonly (n=39) cohorts from the Olink dataset. On day 0, we noted marked upregulation of the angiogenetic factor TNFRSF12A in the infection cohort (q=4.8E-05). Notably, we observed progressive endothelial dysfunction and immune dysregulation in the patients developing severe infections (day 28, Fig. 1B). This included an increased ANG2/1 ratio and dysregulated CD40/CD40L axis. Consistent with a response to microbial exposure, the innate immune regulators CXC3L1, CCL20, and CXCL13 were upregulated in the infection group. Conclusions: The high discriminatory capacity of the HT10 score for infections highlights the advantage of dynamic risk-assessment and emphasizes the utility of procalcitonin in routine inflammatory panels. If validated prospectively, the score will enable more patient-individual decisions on antibiotic use. Finally, the identified Olink candidate markers may further refine the discriminatory capacity of the HT10 score. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Biphasic Neutrophil Recovery after CD19 CART in R/R LBCL Is Associated with Superior PFS/OS, Robust CAR T-Cell Expansion in Relation to Baseline Tumor Volume, and a Decrease of Systemic Inflammation over Time

Background: Hematological toxicity and concomitant infectious complications drive the toxicity burden of CD19 CART. We recently proposed a classification system for CART-related hematotoxicity, defining three unique phenotypes of hematopoietic recovery (Rejeski et al, Blood 2021). The underlying mechanisms of these phenotypes and their relation to survival remain poorly understood. Here, we sought to understand patterns of post-CART neutrophil recovery and their influence on clinical outcomes. Methods: In this multicenter retrospective study, we analyzed 344 pt. receiving standard-of-care axi-cel (n=208) or tisa-cel (n=136) for R/R LBCL. The phenotypes of neutrophil recovery were defined as follows: Quick: sustained neutrophil recovery without a second dip below an ANC<1000/µl.Intermittent: neutrophil recovery (ANC>1500/µl) followed by a second dip with an ANC<1000/µl after day 21.Aplastic: continuous severe neutropenia (ANC<500/µl) ≥14 days. Multivariate analysis of patient-, disease-, and therapy-related factors was performed as binary logistic regression for the aplastic vs. non-aplastic phenotypes. Variables with a p<0.2 on univariate analysis were included. Clinical outcomes were studied via Kaplan-Meier estimates. Quantification of CAR T-cells was performed by qPCR or by flow cytometry in 146 pt. according to institutional protocols. To harmonize CAR T-cell expansion across centers, patients were divided into quartiles by their respective peak expansion. Plasma proteomics were performed in 56 pts. across four time points (day 0, 4, 14, 28) using a 92-protein multiplex proximity extension assay (Olink Proteomics). Key candidate serum markers were validated in 70 pt. using the Ella automated immunoassay system (ProteinSimple). Results: Across all patients, the distribution of the 'aplastic' (A), 'intermittent' (I) and 'quick' (Q) phenotypes was 18%, 42%, and 40%, respectively. The 'aplastic' patients exhibited a longer median duration of severe neutropenia, (A vs. I vs. Q: 26 vs. 10 vs. 5 days, p<0.001), a higher rate of severe infections (A vs. I vs. Q: 41% vs. 23% vs. 18%) and a trend towards higher 1-year non-relapse mortality (A vs. I: 9% vs. 3.5%, p=0.08). On multivariate regression (n=344), the aplastic phenotype was independently associated with the presence of BM infiltration (adjusted OR=3.2, p=0.004), ANC (aOR=0.28, p=0.002), and ferritin (aOR=2.3, p=0.039). Neither high-grade CRS or ICANS nor CAR product were linked to the aplastic phenotype. In the 'intermittent' cohort, we noted a significant decrease of both serum ferritin (median 776 vs. 321 U/L) and CRP levels (median 1.6 vs. 0.2 mg/dL) between CAR infusion and day 90 follow-up. When studying CD3+ CAR T-cell dynamics over time, we found that patients with 'intermittent' recovery displayed both an earlier and more pronounced expansion peak, and longer CART persistence compared to their 'quick' counterparts. While the 'aplastic' cohort displayed comparable peak CART expansion, these patients also exhibited high baseline LDH, indicative of a higher baseline tumor burden, or 'hill to climb' (top right quadrant, Fig. 1A). Conversely, the 'quick' cohort was characterized by both low expansion and low LDH. Serum proteome analysis revealed significant disruptions of endothelial function and immune dysregulation in the 'aplastic' patients at month 1 (Fig. 1B). Importantly, we confirmed higher peak IFNγ and IL-15 levels, and an increased ratio of ANG2 to ANG1 in the 'aplastic' patients in the independent validation cohort. Notably, progression-free (PFS) and overall survival (OS) was poor in the 'aplastic' patients (1-yr PFS 26%, 1-yr OS 46%). On the other hand, 'intermittent' patients exhibited the best survival outcomes (HRPFS I vs. A: 0.45, 95% CI 0.31-0.65). Median PFS was 3 vs. 20 vs. 4 months (A vs. I vs. Q, p<0.001); median OS was 8 months vs. not-reached vs. 23 months (A vs. I vs. Q, p<0.001). Conclusion: The 'aplastic' phenotype is driven by baseline inflammation and hematopoietic function, characterized by progressive endothelial dysfunction, and associated with poor PFS/OS. In contrast, the 'intermittent' phenotype (i.e. biphasic cytopenia) was associated with high levels of CART expansion and persistence, a decrease in systemic inflammation over time, and excellent survival outcomes. Overall, these data suggest potential interactions between host hematopoiesis and CART function and efficacy. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Abrogation of Immune Effector Cell Neurotoxicity Syndrome (ICANS) By Rimiducid (RIM) in Patients Treated with CD19-Specific Chimeric Antigen Receptor Modified T-Cells (CAR-T) Engineered with an Inducible Caspase 9 (iC9 CAR.19)-Clinical and Pharmacodynamic Correlates

Introduction: CD19-targeted CAR-T cells are highly active in B-cell malignancies. While early use of corticosteroids (CS) and tocilizumab have improved the management of cytokine release syndrome (CRS), CS are the mainstay of ICANS management. Though patients with low-grade ICANS often do not require therapy, severe ICANS that does not respond to CS is life-threatening. Rimiducid (RIM) is a dimerizing drug that induces apoptosis in cells engineered with an inducible caspase-9 switch. We reported the first case of the administration of a potentially ablative dose of RIM (0.4 mg/kg) to a patient with B-lymphoblastic leukemia (B-ALL) with severe, prolonged ICANS (Foster MC et al. Blood 2021 Jun 10;137(23):3306-3309). Given this patient's rapid resolution of steroid-nonresponsive ICANS and reduction in iC9 CAR.19 numbers, we sought to explore the effects of lower, potentially non-ablative doses of RIM in patients with CS-nonresponsive ICANS before the development of life threatening complications, with a goal of clinical resolution of ICANS without ablation of CAR T cells. Here, we report the clinical and pharmacodynamic courses of ICANS for the four patients treated with RIM for ICANS to date, in an ongoing expansion cohort of a phase I/II study of iC9 CAR.19 cells in patients with B-ALL. Methods: Adults with B-ALL in 2nd or greater bone marrow (BM) relapse, relapse >100 days after allogeneic stem cell transplant, disease refractory to ≥2 induction therapies, or with measurable residual disease (MRD) persistence/recurrence were enrolled. T-lymphocytes were collected from the subjects, and CAR-T cell products generated by gene modification with a g-retroviral vector encoding for iC9, ΔNGFR (for selection and tracking purposes) and CAR.CD19 (encoding 4-1BB) genes (Diaconu I et al Mol Ther 2017; 25:580). Subjects underwent lymphodepletion with fludarabine and cyclophosphamide followed by infusion of iC9 CAR.19 cells at one of two dose levels (DL1: 5 x 105 CAR-T cells/kg; DL2: 1 x 106 CAR-T cells/kg). Toxicities were graded by CTCAE v5 or ASBMT consensus grading for CRS and ICANS. CAR-T cell expansion in peripheral blood (PB) was measured by flow cytometry (FC) and droplet digital PCR (ddPCR). Cytokines were measured by commercially available multiplex (Luminex®) cytokine assays. Leukemia response was determined by NCCN criteria. Subjects who experienced grade 3-4 ICANS despite CS for >72 hours received RIM 0.4 mg/kg IV over 2 hours. After protocol amendment, subjects with ≥ grade 2 ICANS for >24 hours despite CS received RIM 0.1 mg/kg IV over 2 hours. FC, ddPCR and cytokine levels were measured prior to RIM, at 4 hours after the end of RIM infusion and daily until ICANS resolution. Results: 13 subjects were treated, 3 on DL1 and 10 on DL2, which was the expansion cohort cell dose. Median age was 31 years (range 21-69). CRS developed in 11, with median duration of 5 days (range 1-8). Maximum CRS grade was 2 in 5 and 1 in 6. ICANS developed in 5 subjects, with maximum grade 4 in 2, and grade 3 in 3. One patient with CS-responsive grade 3 ICANS did not meet criteria for RIM. Two subjects with grade 3-4 ICANS for >72 hours received RIM 0.4 mg/kg, both achieving improvement by at least 1 grade within 12 hours and with resolution within 5 days (FIGURE 1). ddPCR for ΔNGFR showed reduction in transcripts by 4 hours after the end of RIM infusion (by 83.6% and 98.6% respectively, FIGURE 2). In two subjects with grade 3 ICANS for 24 hours treated with RIM 0.1 mg/kg, ICANS improved to grade 1 in 24 hours and grade 0 in 48 hours (FIGURE 1). In these subjects, ddPCR for ΔNGFR showed reduction in transcripts at 4 hours after end of RIM infusion by 88.6% and 88.1% respectively (FIGURE 2). Rates of overall response (CR+CRi+MLFS at 4 weeks) were 3/4 RIM subjects versus 9/9 in other subjects. The subject with grade 3 ICANS who did not receive RIM demonstrated peak ΔNGFR transcripts, IL-6, TNF-α and IFNγ two weeks after iC9 CAR.19 infusion, whereas peak transcripts and cytokine levels occurred at one week, and prior to RIM for RIM-treated subjects. Conclusion: RIM administration to patients with ICANS is associated with abrupt reduction of circulating iC9 CAR.19 cells and ICANS grade. Despite detection of CAR-T cells at 3 weeks post infusion and the observance of remissions after treatment with RIM, further investigation of the effects of RIM on iC9 CAR.19 persistence and antileukemic activity is warranted. Lower, potentially less ablative doses of RIM are being explored. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

MRD Status By Clonoseq ® Is a Poor Predictor of Long-Term Outcomes after Bispecific LV20.19 CAR T-Cell Therapy for Relapsed, Refractory B-Cell NHL

INTRODUCTION: The clonoSEQ® Assay (Adaptive Biotechnologies Corporation, Seattle, USA) uses next-generation sequencing (NGS) based testing to identify disease-specific immunoglobulin sequences and detect evidence of minimal residual disease (MRD). MRD assessments have been shown to predict progression-free survival and overall survival in non-Hodgkin's lymphoma (NHL) patients. We report MRD results via the commercially available clonoSEQ® assay and its association with durability of response in patients treated with bispecific LV20.19 CAR in our phase 1/2 trial (NCT04186520). METHODS: We conducted a phase 1/2 single-center prospective trial (NCT04186520) to evaluate the efficacy and safety of LV20.19 CAR T-cells in patients with NHL at 2.5x106 cells/kg dose. LV20.19 CAR T-cells were manufactured onsite at varying lengths of manufacturing (8 vs 12 days) in the CliniMACS Prodigy device with IL7+15 for cell expansion, with the goal of fresh infusion. The commercial clonoSEQ® assay (by Adaptive Biotechnologies) was used to identify and track tumor clonotypes in responding patients at first assessment performed between Day 28-60. We evaluated associations between MRD status and long-term response post LV20.19 CAR-T cell therapy. Descriptive statistics were utilized for baseline characteristics. Survival analyses were calculated utilizing the Kaplan-Meier method and differences between groups were evaluated via the log-rank test. RESULTS: Among the 32 patients enrolled in the trial, there were 26 responders on Day 28. Of these responders, 23 patients had clonoSEQ® B-cell MRD assay done between Day 28-60 post CAR infusion (Table 1). MRD was performed in 11 patients with Diffuse Large B-cell Lymphoma (DLBCL), 3 patients with Follicular lymphoma (FL), and 9 patients with Mantle Cell Lymphoma (MCL). The median age was 64 years (45-74) and the median lines of prior therapy were 4 (2-11). Of the 23 patients with early MRD status available, the day 28 objective response rate (ORR) was 100% (complete response=57%, n=13 and partial response=43%, n=10). Eighteen patients (78%) were MRD negative at first assessment between days 28-60 and 5 (22%) were MRD positive. Of the 5 patients who were MRD positive, 3 became MRD negative at day 90, 1 patient was non-evaluable due to a non-relapse mortality event, and the remaining patient remains positive but with decreasing MRD status at Day 90. None of the patients with MRD positivity at day 28 have relapsed to date. Among patients who achieved early MRD negativity (n=18), 7 (39%) have relapsed, and all were patients with either DLBCL or FL. No MRD-negative MCL patient has relapsed to date. The duration of response in MRD-negative patients was significantly improved in MCL patients compared to DLBCL/FL patients (Figure 1), p=0.02. Among the 7 MRD negative patients who relapsed, only 1 patient was detected to have disease relapse by MRD positivity, 2 were detected simultaneously on MRD testing and imaging, while 3 were detected by physical exam or imaging findings and did not have MRD testing. Interestingly, 1 patient had a central nervous system relapse while being MRD negative. CONCLUSION: Relapse after CAR T-cell therapy remains a significant clinical challenge. New predictors are indicated to determine which responding patients after CAR-T cell therapy are more likely to relapse. Utilizing the commercially available Adaptive clonoSEQ® assay, early MRD assessment was not predictive of long-term clinical outcomes in DLBCL and FL. MRD positive patients maintained durable remission and converted to MRD negativity, while early MRD negative patients continued to relapse. In contrast, MRD negativity post-CAR-T therapy did appear to predict durable clinical remission in MCL patients. Newer assays, including cell-free technologies, may be more informative for non-MCL histologies. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

PO-1174 Radiation therapy as bridging strategy before infusion of CAR T in non hodgkin lymphoma patients

PO-1174 Radiation therapy as bridging strategy before infusion of CAR T in non hodgkin lymphoma patients

Absolute Lymphocyte Count Expansion Kinetics Can be Utilized to Assess Response to CAR T Cell Products

IntroductionCAR T-cell therapies are utilized to treat relapsed/refractory B-lymphoblastic leukemia (B-ALL), diffuse large B cell lymphoma, mantle cell lymphoma, and multiple myeloma. CAR T cell expansion kinetics after infusion impact response. However, these CAR T cell products vary in their target epitope and constituent molecules that makes it difficult to validate broad molecular or flow cytometric assays for use in the clinical setting. The lack of commercially available reagents also limits the ability of clinical laboratories to validate separate CAR T cell specific assays. We investigated the utility of common hematology laboratory parameters to measure CAR T cell expansion and response.MethodsClinical hematology laboratory parameters after infusion of autologous and allogeneic CAR T cell products directed were assessed. Concurrent CBC and cell population data (CPD) parameters from the Sysmex XN 3000 automated hematology analyzer were available in 82 patients. Absolute lymphocyte count (ALC) kinetics after infusion of autologous CD19, allogeneic CD19, CD22, CD33, allogeneic CD123-directed CAR T cell products were analyzed. Patients who received bone marrow transplant served as controls. CPD parameters included X (lateral light scatter-granularity), Y (fluorescence-nuclei acid content), Z (forward scatter-size), and their distribution widths WX, WY, WZ. Archived CellaVision cell morphology images from 118 patients who received CD19-directed CAR T cell products and 25 patients who received other CAR T cell products were analyzed. Response was determined from 1 month post-CAR bone marrow disease assessment.ResultsAbsolute Lymphocyte Counts, lymphocyte morphology and cell population data after infusion of CAR T cell productsCD19-CAR, UCAR19, and CD22-CAR all showed a distinct lymphocyte expansion phase post-infusion in responders (Figure 1A) that was absent in non-responders and controls(Figure 1B). ALC showed characteristic lag, expansion, and contraction phases in responders. CD19-CAR had a peak at day 8 while CD22-CAR and allogenic Universal(U)CAR19 had relatively delayed peak times occurring near day 15 (Figure 1B). The stem cell transplant control patients did not show ALC expansion in the first two weeks and instead showed normal lymphocyte regeneration that occurs in the third to fourth weeks. CD33-CAR and allogeneic UCAR123 non responders did not show ALC expansions.CAR T cell responders showed a distinct sequence of changes in lymphocyte morphology that was absent in non-responders and stem cell transplant controls (Figure 1C). This pattern was noted uniformly across various CAR T cell products (Figure 1D-1F). The morphological changes were categorized as: early, mid, and late. Early atypical cells were noted around days 4-8 after infusion and showed immunoblastic morphology that was present in 89% (n=105) of patients. Mid atypical cells were noted around days 5-14 after infusion and showed atypical large granular lymphocyte morphology that were seen in 95% (n=112) of patients. Late atypical cells showed the typical LGL morphology and was seen in 82% (n=97) of patients.WY fluorescence, which is a measure of nucleic acid content, was most useful in assessing changes in lymphocytes after CAR T cell infusion. WY was low (mean 402, n=57) at baseline pre-infusion timepoint and peaked (mean 1207) approximately 1 week after infusion (mean 7.9 days, n = 59). Peak WY was observed 3.7 days prior to peak ALC (mean day 11.6, n = 59).ConclusionWe demonstrate for the first time that common clinical laboratory parameters can be used to follow CAR T cell expansion after infusion in various autologous and allogeneic CAR T cell products. ALC expansion is a measure of CAR T cell expansion after infusion and correlates with response. Responders showed higher peak ALC compared to non-responders in all CAR T cell products. Timing of peak ALC expansion was determined by other factors such as expression of target antigen and CAR T-cell product used. Lymphocyte morphology followed ALC changes and showed a consistent sequence of changes that was seen across multiple CAR T cell products. Finally, CPD parameter WY which is a measure of nuclei acid content and activation, was an early harbinger of ALC expansion. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Adverse Events Following CAR T-Cell Therapy: A Single Institution Retrospective Analysis of Toxicities Following CAR T-Cell Therapy for Children and Young Adults with Relapsed Refractory B-Cell Acute Lymphoblastic Leukemia

Introduction: CAR T-cells have demonstrated remarkable ability to induce complete remission in patients with relapsed/refractory B-cell acute lymphoblastic leukemia (r/r ALL). This success, however, is tempered by the toxicities associated with CAR T-cell therapy. Much has been published on cytokine release syndrome (CRS), but, to date, a comprehensive profile of specific end organ toxicities secondary to CAR T-cell therapy in the pediatric and young adult population is lacking.Methods: This retrospective, single center study, was performed to characterize the specific adverse events (AEs) experienced by pediatric and young adult patients during the first 30 days following CAR T-cell infusion. AEs graded using Common Terminology Criteria for Adverse Events (CTCAE) were collected from all patients with r/r ALL treated on one of three phase I CAR T-cell trials (CD19, CD22, and CD1922) at the Pediatric Oncology Branch of the National Cancer Institute at the National Institutes of Health from 2012-2020. The primary objective was to determine the incidence of all severe AEs, defined as > grade (Gr) 3 AEs, overall and by organ system, attributed to research or disease. Secondary objectives were to stratify severe AEs based on development of CRS and CRS grade (using ASTCT CRS grading criteria). Descriptive statistics were reported along with comparisons of continuous parameters using Mann-Whitney and binomial parameters using Fischer's exact tests.Results: We reviewed AE data from 134 patients with r/r ALL receiving one of 3 unique CAR T-cell constructs (Table). The median age was 15.2 years (Interquartile range (IQR) 9.5-21.2). The median number of prior therapies was 5 (IQR 3-6) and 57% had received a prior hematopoietic stem cell transplant (HSCT).Amongst the 134 patients, a total of 1747 individual > Gr 3 AEs were experienced by 133 patients (99%) during the first 30 days following CAR infusion (Figure 1A). The median number of > Gr 3 AEs per patient was 10 (IQR 4.8-19).Cytopenias (including neutropenia, thrombocytopenia and anemia) comprised the vast majority of total > Gr 3 AEs (n=983, 56.3%). The most common severe (> Gr 3) AEs were thrombocytopenia (n=433, 24.8%), metabolic abnormalities (i.e. electrolyte derangements) (n=333, 19.1%), neutropenia (n=332, 19%), and anemia (n=218, 12.5%).With exclusion of cytopenias, 764 > Gr 3 AEs were experienced by 111 patients (83%), with a median of 4 (IQR 1-8.3) > Gr 3 AEs per patient. One grade 5 pulmonary AE occurred in the setting of acute respiratory distress syndrome (ARDS). Focusing on non-cytopenia AEs (Figure 1B), metabolic AEs made up 43.6% of AEs; hepatic toxicities (n=115, 15%), febrile neutropenia (n=114, 14.9%), and cardiovascular toxicities (n=59, 7.7%) were the next most frequent.Of the 134 patients, 104 (77.6%) developed CRS. All 30 patients without CRS had at least 1 > Gr 3 AE (median 6, IQR 3-11.3). In contrast, the median number of > Gr 3 AEs in those with CRS was 11.5 (IQR, 6-21.5), (p=0.0021). When stratified by CRS Gr 1-2 versus CRS Gr 3-4 (Figure 2), patients with higher-grade CRS also had a higher median number of > Gr 3 AEs per patient (p= 0.0017).Conclusions: Among 134 children and young adults with r/r ALL receiving phase I CAR T-cells, we found a high incidence (99%) of severe AEs, with a per patient median of 10 (IQR 4.8-19) > Gr 3 AEs. While the majority of > Gr 3 AEs were cytopenias, 17 different categories of AEs were experienced. The development and severity of CRS associated with an increase in the median number of severe AEs per patient. As phase I trials of CAR T-cell therapy expand, it is imperative to understand the full toxicity profile of these therapies. While the definition and refined grading of CRS has helped advance the field, there is a gap in knowledge regarding patient specific end-organ toxicities beyond CRS. Our data help establish a foundation for the full toxicity profile experienced by patients enrolling on phase I CAR T-cell trials. With an emerging role for earlier intervention for CRS, we anticipate that the toxicity burden will decrease. Next steps include characterizing the specific toxicities within each AE category, evaluating duration and time to resolution, distinguishing attribution to research versus disease and studying the impact of earlier use of tocilizumab on toxicity profile. Future directions will incorporate assessment of baseline organ function pre-CAR and its impact on development of post CAR severe AEs. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Relmacabtagene Autoleucel CD19 CART Therapy for Adults with Heavily-Pretreated Relapsed/Refractory Large B-Cell Lymphoma in China

IntroductionPrognosis for relapsed and refractory large B-cell lymphoma (r/r LBCL) is poor, and treatment remains challenging. Relmacabtagene autoleucel (relma-cel) is a CD19-directed 4-1BB/CD3z chimeric antigen receptor T cell (CAR-T) product manufactured in China. RELIANCE study is the first CAR-T study with CD19 as target that got IND approved by the authority in China. Initial findings of the pivotal Phase 2 single-arm, multicenter RELIANCE trial demonstrated high response rates and low rates of CAR-T-associated toxicity with relma-cel treatment in heavily pretreated patients (pts) with r/r LBCL (NCT04089215; Ying et al. Cancer Med 2020;10:999-1011). Long-term follow-up data for the RELIANCE study are presented.Materials & MethodsFifty-nine adults (age ≥18 years) with heavily pretreated (≥2 prior therapies), measurable and histologically confirmed r/r LBCL were randomized to receive lymphodepletion chemotherapy (LDC: fludarabine 25 mg/m 2 + cyclophosphamide 250 mg/m 2 daily×3) followed by a single infusion of autologous CAR+ T cells at a dose of 100×10 6 or 150×10 6. Pts were evaluated for efficacy using Lugano criteria and for toxicity using Common Terminology Criteria for Adverse Events v4.03 and for cytokine release syndrome (CRS) using Lee et al. (Blood 2014;124(2):188-95). Primary endpoint was 3-month objective response rate (ORR); key secondary endpoints included complete response rate (CRR) at 3 months, duration of response (DOR), progression-free survival (PFS), overall survival (OS) and treatment-emergent adverse event (TEAE) profile. The data cutoff date was December 31, 2020.ResultsAt the time of analysis, all 59 pts (median age 56.0 years, range 18-75 years) were at least 15 months post-treatment with relma-cel (7 pts completed trial, 34 pts on study, 18 pts withdrew). Among the modified intent-to-treat (mITT) population of 58 efficacy-evaluable pts, best ORR was 77.6%, with a best CRR of 51.7%; ORR and CRR at 3-month-postdose landmark were 60.3% and 44.8%, respectively. With a median follow-up of 17.9 months (range 0.3-25.6 months), median PFS (mITT) was 7.0 months (95% CI 4.8-not reached [NR]). Median PFS was NR (95% CI 8.8 months-NR) for pts with compete response (CR) at 3 months, the 6-, 12-, 18- and 24-month PFS rates were 80.8%, 69.2%, 69.2% and 69.2%, respectively (Figure). PFS was associated with objective response or CR at 1, 3, 6, and 12 months (log-rank test, p≤0.002). Median OS was NR (95% CI NR-NR) for both the mITT population and pts with CR at 3 months (12-month OS rate, 76.8% and 92.3%, respectively). Median DOR was NR (95% CI 4.9 months-NR) for the mITT population and (95% CI 8.0 months-NR) for pts with CR at 3 months. Levels of CAR-T cells declined to below the level of quantification (BLQ) in 41 (69.5%) pts, among whom nearly 40% had sustained response. Median time from documentation of CAR-T BLQ to disease progression was 6.1 months (95% CI 1.8-NR). Treatment-related TEAEs were reported in 93.2% of pts and were primarily grade 1-2 (89.8% pts); 54.2% of pts had grade ≥3 TEAEs. Pyrexia was the most common TEAE, reported in 59.3% of pts (all grade 1-2). The most common grade ≥3 treatment-related TEAEs were neutrophil count decrease (30.5%) and white blood cell (WBC) count decrease (13.6%). Presence of cytopenia at day 29 (78.0%) was significantly associated with WBC count after LDC, change in serum albumin between days 1 and 4 and serum interleukin 8 after LDC (p<0.05). The incidence of CRS and neurotoxicity (NT) was 47.5% and 20.3%, respectively, both primarily grade 1-2. There were no CRS- or NT-related deaths. Occurrence of grade ≥2 CRS or NT (11.9% and 6.8%, respectively) was significantly associated with fold-change of platelet count on day 4 compared with baseline and day 1 absolute neutrophil count (p<0.03). There were no treatment-related deaths.ConclusionsThese are the first data of long-term follow-up CD19 CAR-T study with IND approval in China reported. With nearly 18 months of median follow-up, the RELIANCE pivotal registered study demonstrated durable responses, high rates of OS and PFS, and no new safety profile found in r/r LBCL pts. Based on these data, relma-cel is undergoing approval in China. Figure: Kaplan-Meier estimates of (A) PFS and (B) OS by 3-month tumor responsePR, partial response [Display omitted] DisclosuresZhang: JW Therapeutics: Current Employment. Yang: JW Therapeutics: Current Employment. Zhou: JW Therapeutics: Current Employment. Zheng: JW Therapeutics: Current Employment.

Outcomes after Reinfusion of CD19-Specific Chimeric Antigen Receptor (CAR)-Modified T Cells in Children and Young Adults with Relapsed/Refractory B-Cell Acute Lymphoblastic Leukemia

Background: CAR-modified T cells targeting CD19 have produced remarkable responses in relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL); however, relapse continues to be a substantial challenge. CD19+ relapses, which account for 33-78% of relapses, are associated with loss of CAR T-cell surveillance due to short persistence. Thus, strategies to improve functional persistence to prevent and treat CD19+ relapsed disease are crucial. Here, we report our experience administering reinfusions of murine or humanized 4-1BB CD19 CAR T cells in an effort to prolong persistence in patients with demonstrated short persistence to mitigate relapse risk, treat CD19+ relapsed disease, and produce responses after nonresponse to initial CAR infusion.Methods: This analysis included patients aged <30 years treated with a murine CD19 CAR construct, either investigational CTL019 (NCT01626495, NCT02906371) or commercial tisagenlecleucel, or a humanized CD19 CAR construct, huCART19 (NCT02374333), who received ≥1 reinfusion of the same CAR product due to: 1) clinical signs of poor persistence within 6 months (mos) of initial infusion, including peripheral B-cell recovery (BCR) or CD19+ hematogones in the bone marrow, 2) new CD19+ minimal residual disease (MRD) or relapse, or 3) nonresponse to initial infusion. The huCART19 trial included patients who had previously received a CAR T cell product (CAR-exposed), whereas all patients reinfused with CTL019/tisagenlecleucel were CAR-naïve at initial infusion. The primary outcome was complete response (CR) at day 28 after reinfusion, defined as complete remission with establishment or maintenance of B-cell aplasia. Secondary outcomes included CRS incidence, cumulative incidence of relapse (CIR) and overall survival (OS).Results: Among 229 CAR-naïve and 33 CAR-exposed patients treated with CD19 CAR between 2012-2020, 81 received ≥1 reinfusion (investigational CTL019, n=44; commercial tisagenlecleucel, n=11; huCART19, n=13 CAR-naïve and n=13 CAR-exposed). In addition, 18 patients received PD-1 blockade after their first (n=11) or subsequent (n=7) reinfusions. Indications for first reinfusion were peripheral BCR (CAR-naïve, n=32; CAR-exposed, n=6), hematogones (CAR-naïve, n=21; CAR-exposed, n=4), CD19+ MRD/relapse (CAR-naïve, n=10, CAR-exposed, n=0), and nonresponse to initial infusion (CAR-naïve, n=5, CAR-exposed, n=3). CRS grade ≥2 (Penn scale) occurred in 19 patients (grade 2, n=13; grade 3, n=4; grade 4, n=2). Grade 3-4 events only occurred in patients with active disease at time of reinfusion. Twenty-two patients had an inpatient admission within 30 days of first reinfusion, of which 7 required intensive care unit admissionAmong the 63 patients reinfused for relapse prevention, 33 (52%) had a CR at day 28. With a median duration of follow-up of 38 mos, 13 experienced a subsequent relapse (7 CD19+, 4 CD19-, 2 CD19-subset negative), 4 received alternative therapy or allogeneic hematopoietic stem cell transplantation (HSCT) in remission, and 16 remain in remission without further therapy at a median of 39 mos after first reinfusion. The median duration of B-cell aplasia was 8 mos (IQR 2-35) after reinfusion. Of the 30 with no response (NR), 10 had a subsequent CD19+ relapse, 15 received alternative therapy or HSCT, and 5 remain in remission without further therapy at a median of 43 mos after reinfusion. CIR and OS were not statistically significantly different between patients with CR or NR (CIR, p=0.26; OS, p=0.25) (Figure A-B). However, at 24 mos after reinfusion, CIR was 29% (95% CI, 11-44%) for CR compared to 61% (95% CI, 24-80%) for NR; OS was 90% (95% CI, 80-100%) for both groups.Of the 10 patients reinfused for relapse, 5 (50%) had a CR; 2 subsequently experienced a CD19+ relapse, 2 received an HSCT in remission, and 1 remains in remission without further therapy at 18 mos after reinfusion. Of the 8 patients reinfused for nonresponse to initial infusion, 7 were evaluable; none had a CR, and all died at a median of 2.5 mos after reinfusion.Conclusions: Reinfusion of CTL019/tisagenlecleucel or huCART19 is safe, may prolong B-cell aplasia in patients with short CAR persistence and reduce relapse risk, and can induce remissions in patients with CD19+ relapsed disease. Thus, reinfusion may provide an alternative to HSCT for short persistence. However, reinfusion is not effective for patients with nonresponse to initial CAR infusion. [Display omitted] DisclosuresCallahan: Novartis: Speakers Bureau. Rheingold: Optinose: Other: Spouse's current employment; Pfizer: Research Funding. June: Tmunity, DeCART, BluesphereBio, Carisma, Cellares, Celldex, Cabaletta, Poseida, Verismo, Ziopharm: Current equity holder in publicly-traded company; Novartis: Patents & Royalties; AC Immune, DeCART, BluesphereBio, Carisma, Cellares, Celldex, Cabaletta, Poseida, Verismo, Ziopharm: Consultancy. Grupp: Novartis, Roche, GSK, Humanigen, CBMG, Eureka, and Janssen/JnJ: Consultancy; Novartis, Adaptimmune, TCR2, Cellectis, Juno, Vertex, Allogene and Cabaletta: Other: Study steering committees or scientific advisory boards; Novartis, Kite, Vertex, and Servier: Research Funding; Jazz Pharmaceuticals: Consultancy, Other: Steering committee, Research Funding. Maude: Novartis Pharmaceuticals Corporation: Consultancy, Research Funding; Wugen: Consultancy.

Cytokine release syndrome after CAR infusion in pediatric patients with refractory/relapsed B-ALL: is there a role for diclofenac?

Cytokine release syndrome (CRS) is a major complication after chimeric-antigen receptor T-cell treatment, characterized by an uncontrolled systemic inflammatory reaction. We investigated the potential role of diclofenac in the management of CRS in five pediatric patients treated for relapsed/refractory B-lineage acute lymphoblastic leukemia. In case of persistent fever with fever-free intervals shorter than 3 hours, diclofenac continuous infusion was initiated, at the starting dose of 0.5 mg/Kg/day, the lowest effective pediatric dose in our experience, possibly escalated up to 1 mg/Kg/day, as per institutional guidelines. CRS occurred at a median of 20 hours (range 8-27) after tisagenlecleucel infusion. Diclofenac was started at a median of 20 hours (range 13-33) after fever onset. A mean of 3.07 febrile peaks without diclofenac and 0.95 with diclofenac were reported (p = 0.02). Clinical benefit was achieved by hampering the progression of tachypnea and tachycardia. Despite fever control, CRS progressed in four of the five patients, and hypotension requiring vasopressors and fluid retention, as well as hypoxia, occurred. Vasopressors were followed by 1-2 doses of tocilizumab (one in patient 2 and two in patients 3, 4, and 5), plus steroids in patients 4 and 5. Based on a limited number of patients, diclofenac leads to better fever control, which translates into symptom relief and improvement of tachycardia, but could not prevent the progression of CRS.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature